CN114163539A - TP recombinant antigen and preparation method and application thereof - Google Patents

TP recombinant antigen and preparation method and application thereof Download PDF

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CN114163539A
CN114163539A CN202111523961.7A CN202111523961A CN114163539A CN 114163539 A CN114163539 A CN 114163539A CN 202111523961 A CN202111523961 A CN 202111523961A CN 114163539 A CN114163539 A CN 114163539A
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杨帆
李林
刘万建
刘洋
李文
王婷
田永帅
宋金玲
高文晓
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Qingdao Shuojing Biotechnology Co ltd
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Abstract

The invention belongs to the technical field of immunodetection, and relates to a TP recombinant antigen and a preparation method and application thereof, wherein the TP recombinant antigen comprises at least 1 TP17 antigen, at least 1 TP15 antigen and at least 1 TP47 antigen; the amino acid sequence of the TP17 antigen is shown as SEQ ID NO: 1-2; the amino acid sequence of the TP15 antigen is shown as SEQ ID NO: 3 is shown in the specification; the amino acid sequence of the TP47 antigen is shown as SEQ ID NO: 4-6. The TP recombinant antigen is based on TP17, TP15 and TP47 outer membrane proteins with strong immunogenicity, effective antigen epitopes are selected, and the effective antigen epitopes can be fully and effectively exposed by carrying out combined expression and different sequence arrangement on the effective antigen epitopes, so that the missed detection is prevented; the mutation of cysteine solves the false positive problem caused by protein aggregation, so that the protein has obvious advantages in the aspect of antigen specificity; the application specificity and the sensitivity are obviously improved.

Description

TP recombinant antigen and preparation method and application thereof
Technical Field
The invention belongs to the technical field of genetic engineering and immunodetection, and particularly relates to a TP recombinant antigen and a preparation method and application thereof.
Background
Syphilis (Syphilis) is a chronic, systemically transmitted disease caused by treponema pallidum, which is found in both the skin, secretions, and blood of dominant and recessive Syphilis patients. Treponema Pallidum (TP), also known as Treponemiapallidum, is the causative agent of venereal disease, syphilis, and the TP gene is circular DNA composed of 1138006bp, with 1041 open reading frames, and 55% of the reading frame translation proteins have biological functions. The treponema pallidum outer membrane does not contain lipopolysaccharide, but contains abundant lipoprotein, and the TP is supposed to have 22 kinds of lipoprotein, and the contents of the TP are more Tp47, Tp44.5, Tp36, Tp35, Tp17, Tp15 and the like. Wherein, the outer membrane lipoproteins Tp47, Tp17, Tp15, Tp44.5 and the like have high immunogenicity and have important significance in the immunodiagnosis of the syphilis.
The gene coding TP17(17kDa protein) is Tpp17, the length of the gene is 468bp, 156 amino acids are coded, the N end of the protein is provided with cysteine residues modified by fatty acid, a hydrophobicity experiment shows that a lipid modification structure is related to the hydrophobicity of TP17, further research proves that the lipid modification structure can stimulate macrophages of mice to express tumor necrosis factors, is related to pathogenicity and possibly plays an important role in syphilis pathogenesis, and meanwhile, the TP17 has multiple antigenic determinants and strong immunogenicity, and almost all TP infectors can generate anti-TP 17 antibodies with high titer specificity. The gene encoding TP15(15KDa protein) is Tpp15, the gene length is 426bp, and TP15 lipoprotein is a main immunogen for natural TP infection and experimental TP infection, and can cause strong humoral immunity, cellular immunity and resistance to TP reinfection at the later stage of infection in a rabbit immune model, so that the gene is considered to play an important role in protective immunity. The gene coding TP47(47kDa protein) is Tpp47, and the length of the gene is 1302 bp. The TP47 protein is an integral membrane protein and is one of the highest-content components in the TP outer membrane protein; TP47 has multiple biological functions, has penicillin binding activity, is a zinc ion-dependent carboxypeptidase, and has strong immunogenicity. The three protein gene sequences are widely applied to diagnosis and vaccine research due to high conservation and high immunogenicity.
With the development of genetic engineering technology, molecular biological research on TP gene is becoming mature and applied to diagnostic reagents. Antibodies against TP15, TP17, TP47 and the like can be detected at each stage of the syphilis course, and the antibodies account for absolute dominance in the serum of patients in the advanced stage of latent infection, so the TP diagnostic reagent mainly uses gene fragments of TP17, TP15, TP47 and the like on different detection platforms, but the TP diagnostic raw materials in the diagnostic reagent are mostly in a mode of single TP17, single TP47, TP17+ TP15 chimeric, TP17+ TP47 chimeric, TP15+ TP47 chimeric and the like. Due to the diversity of clinical samples and the differences in the time and amount of antibody production of different segments, the chimerism of a single fragment or two fragments is likely to result in missed clinical tests, and the selection of different fragments of the protein also results in cross-reactivity and thus false positive results.
Thus, there remains a need in the art for products that avoid the omission of clinical testing and detect TP with high sensitivity and specificity for rapid, accurate, comprehensive screening and diagnosis.
Specific detection methods using treponema pallidum as an antigen include a treponema pallidum fluorescent antibody adsorption test (FTA-ABS) and a treponema pallidum hemagglutination Test (TPHA), and the specificity and the sensitivity are high. However, because of the difficulty in culturing treponema pallidum, the cost of the reagent is high, and because the treponema pallidum antigen is complex, the treponema pallidum antigen can still have a certain nonspecific reaction with other pathogens, especially similar spirochaete diseases (such as yasis).
In view of the above, scientists have begun to use recombinant proteins as specific detection methods for antigens. With the development of gene cloning technology, immunoassay methods using recombinant syphilis proteins as antigens have been applied successively, and currently, treponema pallidum serum immunological detection methods mainly utilize enzyme-linked immunosorbent assay, chemiluminescence assay, gold-labeled chromatography and the like based on the principle of a double-antigen sandwich method.
The principle of the TP double antigen sandwich method is as follows: coating a first antigen (namely a coating antigen) on a solid phase support, adding a sample to be detected, introducing a second antigen (namely a labeling antigen) labeled with a special label in a proper mode, forming a sandwich structure of coating antigen-TP antibody-labeling antigen by using TP antibody and the two antigens if the TP antibody exists in the sample, and amplifying a signal by using the label on the labeling antigen to obtain a final judgment result. In terms of methodology, the current syphilis ELISA and colloidal gold detection methods are double-antigen sandwich methods directly labeled by a label and a labeled antigen, but the double-antigen sandwich methods have the following defects: when the marker is small markers such as enzyme, luminescent substance and the like, the marking proportion needs to be increased for improving the sensitivity, but the marked antigen is wrapped by the marker due to too high proportion, so that the antigen epitope is embedded to cause the reduction of the antigen activity, finally the sensitivity is low, and the phenomenon of omission occurs; when the marker is a large nanoparticle such as colloidal gold, latex or other nanoparticle markers, the labeled antigen is too low and is easy to precipitate, so that the sensitivity is low, the specificity is reduced due to too high labeled antigen, and meanwhile, the antigen activity is reduced due to the fact that the labeled antigen epitope is embedded due to too large marker, so that further improvement on the existing syphilis double-antigen sandwich method kit is urgently needed at present, the sensitivity of the kit is improved, and the specificity is improved at the same time.
Disclosure of Invention
The invention aims to solve the problems in the prior art, provides a TP recombinant antigen, a preparation method and application thereof, and also provides nucleic acid for coding the TP recombinant antigen and a mutant thereof, and a related vector, a host cell, an immunoassay method, a detection kit and the like. The TP recombinant antigen and the product thereof provided by the invention can be applied to detecting the existence of TP antibody in a sample from a subject, and compared with the existing detection method, the sensitivity and specificity of the detection provided by the invention are obviously improved.
The technical scheme of the invention is as follows:
a TP recombinant antigen comprising at least 1 TP17 antigen, at least 1 TP15 antigen, and at least 1 TP47 antigen;
wherein, the TP17 antigen amino acid sequence is selected from GeneBank: WP _010881883.1 sequence polypeptide selected from polypeptide 23-156aa in length, 23-145aa in which the 42 th cysteine and the 58 th cysteine are mutated to alanine, i.e. C42A, C58A; the amino acid sequence of the TP17 antigen is as follows:
VSCTTVCPHAGKAKAEKVEAALKGGIFRGTLPAADAPGIDTTVTFNADGTAQKVELALEKKSAPSPLTYRGTWMVREDGIVELSLVSSEQSKAPHEKELYELIDSNSVRYMGAPGAGKPSKEMAPFYVLKKTKK(SEQ ID NO：1)
or, VSCTTVCPHAGKAKAEKVEAALKGGIFRGTLPAADAPGIDTTVTFNADGTAQKVELALEKKSAPSPLTYRGTWMVREDGIVELSLVSSEQSKAPHEKELYELIDSNSVRYMGAPGAGKPSKEM (SEQ ID NO: 2)
Wherein, the TP15 antigen amino acid sequence is selected from GeneBank: an AAC65160.1 sequence polypeptide, the selected polypeptide being 22-141aa in length; the amino acid sequence of the TP15 antigen is as follows:
SSIPNGTYRATYQDFDENGWKDFLEVTFDGGKMVQVVYDYQHKEGRFKSQDADYHRVMYASSGIGPEKAFRELADALLEKGNPEMVDVVTGATVSSQSFRRLGAALLQSARRGEKEAIISR(SEQ ID NO：3)
wherein, the TP47 antigen amino acid sequence is selected from GeneBank: AAC65545.1 sequence polypeptide, the length of the selected polypeptide can be 68-410aa, 239-419aa and 21-434aa, wherein the 239 th amino acid glycine is mutated into valine, namely G239V; the amino acid sequence of the TP47 antigen is as follows:
MFDAVSRATHGHGAFRQQFQYAVEVLGEKVLSKQETEDSRGRKKWEYETDPSVTKMVRASASFQDLGEDGEIKFEAVEGAVALADRASSFMVDSEEYKITNVKVHGMKFVPVAVPHELKGIAKEKFHFVEDSRVTENTNGLKTMLTEDSFSARKVSSMESPHDLVVDTVGTVYHSRFGSDAEASVMLKRADGSELSHREFIDYVMNFNTVRYDYYGDDASYTNLMASYGTKHSADSWWKTGRVPRISCGINYGFDRFKGSGPGYYRLTLIANGYRDVVADVRFLPKYEGNIDIGLKGKVLTIGGADAETLMDAAVDVFADGQPKLVSDQAVSLGQNVLSADFT(SEQ ID NO：4)
or, VYHSRFGSDAEASVMLKRADGSELSHREFIDYVMNFNTVRYDYYGDDASYTNLMASYGTKHSADSWWKTGRVPRISCGINYGFDRFKGSGPGYYRLTLIANGYRDVVADVRFLPKYEGNIDIGLKGKVLTIGGADAETLMDAAVDVFADGQPKLVSDQAVSLGQNVLSADFTPGTEYTVEV (SEQ ID NO: 5)
Or, GSSHHETHYGYATLSYADYWAGELGQSRDVLLAGNAEADRAGDLDAGMFDAVSRATHGHGAFRQQFQYAVEVLGEKVLSKQETEDSRGRKKWEYETDPSVTKMVRASASFQDLGEDGEIKFEAVEGAVALADRASSFMVDSEEYKITNVKVHGMKFVPVAVPHELKGIAKEKFHFVEDSRVTENTNGLKTMLTEDSFSARKVSSMESPHDLVVDTVGTVYHSRFGSDAEASVMLKRADGSELSHREFIDYVMNFNTVRYDYYGDDASYTNLMASYGTKHSADSWWKTGRVPRISCGINYGFDRFKGSGPGYYRLTLIANGYRDVVADVRFLPKYEGNIDIGLKGKVLTIGGADAETLMDAAVDVFADGQPKLVSDQAVSLGQNVLSADFTPGTEYTVEVRFKEFGSVRAKVVAQ (SEQ ID NO: 6)
The segments selected for the TP17 antigen and the segments selected for the TP47 antigen contained in the TP recombinant antigen may be the same or different.
Further, the TP recombinant antigen comprises 1 TP17 antigen, 1 TP15 antigen and 1 TP47 antigen.
Further, the TP recombinant antigen comprises 2TP 17 antigens, 1 TP15 antigen and 1 TP47 antigen; wherein, the amino acid sequences of 2TP 17 antigens are all shown in SEQ ID NO: 1 or 2 of the amino acid sequences of the TP17 antigens are all shown as SEQ ID NO: 2, or 2 amino acid sequences of the TP17 antigens are shown as SEQ ID NOs: 1 and SEQ ID NO: 2, or a pharmaceutically acceptable salt thereof.
Further, the TP17 antigen, the TP15 antigen, and the TP47 antigen are directly linked or optionally linked through one or more linkers, which are linking peptides, including (G) n, (GS) n, (SG) n, (GGGS) n, (GGGGS) n, or (AAY) n, wherein n is an integer of 1, 2, 3, 4, 5, or more.
Further, the antigen contained in the antigen is TP17-TP15-TP47, or TP15-TP17-TP47, or TP17-TP15-TP47-TP17, or TP17-TP17-TP15-TP47 in the sequence from the N terminal to the C terminal.
Furthermore, the TP recombinant antigen can be a labeled antigen or a coating antigen; the N end of the TP recombinant antigen serving as a labeled antigen comprises a TRX label, and the N end of the TP recombinant antigen serving as a coating antigen comprises a GST label.
The invention also protects a nucleic acid encoding the TP recombinant antigen;
further, an expression vector comprising the above nucleic acid is also protected; and host cells, such as E.coli cells, comprising the above expression vectors.
The invention also provides a syphilis detection test strip which comprises a coating antigen, a labeled antigen, a TRX monoclonal antibody and a marker; the labeled antigen is a TP recombinant antigen of which the N end contains a TRX label, and the coating antigen is a TP recombinant antigen of which the N end contains a GST label; the label is indirectly bound to the labeled antigen through the TRX monoclonal antibody.
The invention has the beneficial effects that:
(1) the TP recombinant antigen provided by the invention is based on TP17, TP15 and TP47 outer membrane proteins with strong immunogenicity, selects effective antigen epitopes, performs combined expression on the effective antigen epitopes, and performs different sequence arrangement, so that the antigen epitopes can be fully and effectively exposed, and the omission is prevented; meanwhile, aggregation caused by disulfide bonds is avoided by mutation of cysteine, so that the problem of false positive caused by protein aggregation is solved, and the protein has obvious advantages in the aspect of antigen specificity. The protein is respectively used as a coating antigen and a labeled antigen to be applied to a gold-labeled chromatography platform to detect TP antibodies, so that the specificity and the sensitivity of detection are obviously improved, and the development of a high-quality colloidal gold reagent is facilitated.
(2) In order to solve the problem of low clinical TP antibody detection sensitivity, the invention constructs brand-new TP genetic engineering recombinant antigen protein through combination and chimeric modes of different fragments and a large amount of grope, and the prepared TP recombinant antigen effectively improves the detection sensitivity of the antibodies against TP17, TP15 and TP47 and avoids the problem of clinical missed detection;
meanwhile, in order to further improve the specificity of clinical TP antibody detection, mutation treatment is carried out on partial amino acids of TP17 fragments in a TP recombinant antigen, different fragment screening is carried out on TP47 in order to avoid cross reaction, the specificity of clinical TP antibody detection is effectively improved, and the problem of false positive is reduced.
(3) In order to further improve the labeling effect, the invention introduces the monoclonal antibody aiming at the TRX label, improves the labeling efficiency, reduces the dosage of the labeled antigen and improves the clinical detection sensitivity of the TP antibody by an indirect labeling method.
Drawings
FIG. 1 is a flow chart of a recombinant antigen method provided by the present invention;
FIG. 2 is a schematic structural diagram of a colloidal gold test strip provided by the present invention;
FIG. 3 is a schematic diagram illustrating the results of the colloidal gold test strip provided by the present invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
For a further understanding of the invention, reference will now be made to the following description taken in conjunction with the accompanying drawings and examples.
EXAMPLE 1 construction of plasmid vector for recombinant protein in Gene engineering
Designing a gene segment of TP recombinant protein, wherein TP17 refers to GenBank database WP _010881883.1 amino acid sequence, and the length of the selected polypeptide can be 23-156aa (TP17-1) and can also be 23-100aa (TP 17-2); wherein the cysteine at position 42 and cysteine at position 58 are mutated to alanine, i.e., C42A, C58A; synthesis of TP17-1 nucleic acid sequence by codon optimization SEQ ID NO: 7.
TP15 amino acid sequence reference GeneBank: AAC65160.1 amino acid sequence, the selected polypeptide being 22-141aa in length; synthesis of TP15 nucleic acid sequence by codon optimization SEQ ID NO: 8.
the TP47 antigen amino acid sequence is selected from GeneBank: AAC65545.1 sequence polypeptide, the selected polypeptide may be 68-410aa (TP47-1), 239-419aa (TP47-2), 21-434aa (TP47-3), the nucleotide sequence of TP47-3 is synthesized by codon optimization, SEQ ID NO: 9.
the nucleic acid sequence of SEQ ID NO: 7-9 were all synthesized by Suzhou Hongxn Biotechnology Ltd.
The gene segments provided by the embodiment are spliced in a mode of a primer, namely, a double bag by nested PCR, and are finally constructed on a PET32a expression vector in a mode of enzyme digestion and connection by restriction enzyme and T4 DNA ligase, and a TP-1 recombinant antigen is constructed on a pGEX-4T-1 expression vector at the same time.
The following recombinant TP antigens were prepared according to the above description, and each recombinant TP antigen was fused in the order from N-terminus to C-terminus, and the antigens were linked directly or via one or more linker peptide linkers to prepare the following recombinant antigens:
TP-1 recombinant antigen fused with TP17-1 segment, TP15 segment and TP47-3 segment from N terminal to C terminal, wherein the connecting peptide between TP17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-3 is GSG3SGSGSG; the nucleotide sequence of the prepared TP-1 recombinant antigen is shown in SEQ ID NO: 10, the amino acid sequence is shown in SEQ ID NO: 11.
TP-2 recombinant antigen fused with TP17-1 segment, TP15 segment and TP47-2 segment from N terminal to C terminal, wherein the connecting peptide between TP17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-2 is GSG3SGSGSG; the nucleotide sequence of the prepared TP-2 recombinant antigen is shown in SEQ ID NO: 12, the amino acid sequence is shown in SEQ ID NO: 13.
TP-3 recombinant antigen fused with TP17-1 fragment, TP15 fragment and TP47-1 fragment from N terminal to C terminal, wherein the TP17-1 and the TP15 are connected with each otherThe connecting peptide is (GGGGS)2, and the connecting peptide between TP15 and TP47-1 is GSG3SGSGSG; the nucleotide sequence of the prepared TP-3 recombinant antigen is shown in SEQ ID NO: 14, amino acid sequence shown in SEQ ID NO: 15.
TP-4 recombinant antigen, which is fused with a TP15 fragment, a TP17-1 fragment and a TP47-2 fragment from N end to C end, wherein the connecting peptide between TP15 and TP17-1 is (AAY)2, and the connecting peptide between TP17-1 and TP47-2 is (GGGGS) 3; the nucleotide sequence of the prepared TP-4 recombinant antigen is shown in SEQ ID NO: 16, amino acid sequence shown in SEQ ID NO: 17.
TP-5 recombinant antigen, which is fused with a TP15 fragment, a TP17-1 fragment and a TP47-3 fragment from an N end to a C end, wherein a connecting peptide between TP15 and TP17-1 is (AAY)2, and a connecting peptide between TP17-1 and TP47-3 is (GGGGS) 3; the nucleotide sequence of the prepared TP-5 recombinant antigen is shown in SEQ ID NO: 18, amino acid sequence shown in SEQ ID NO: 19.
TP-6 recombinant antigen fused with TP17-1 segment, TP17-2 segment, TP15 and TP47-2 segment from N terminal to C terminal, wherein the connecting peptide between TP17-1 and TP17-1 is (AAY)2, the connecting peptide between TP17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-3 is GSG3SGSGSG; the nucleotide sequence of the prepared TP-6 recombinant antigen is shown in SEQ ID NO: 20, the amino acid sequence is shown in SEQ ID NO: 21.
TP-7 recombinant antigen fused with TP17-1 segment, TP17-1 segment, TP15 segment and TP47-2 segment from N terminal to C terminal, wherein the connecting peptide between TP17-1 and TP17-1 is (AAY)2, the connecting peptide between TP17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-2 is GSG3SGSGSG; the nucleotide sequence of the prepared TP-7 recombinant antigen is shown in SEQ ID NO: 22, amino acid sequence shown in SEQ ID NO: 23.
TP-8 recombinant antigen fused with TP17-1 segment, TP15 segment, TP47-3 segment and TP17-1 segment from N terminal to C terminal, wherein the connecting peptide between TP17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-3 is GSG3The connecting peptide between SGSGSG, TP47-3 and TP17-1 is (AAY) 2; the nucleotide sequence of the prepared TP-8 recombinant antigen is shown in SEQ ID NO: 24, the amino acid sequence is shown in SEQ ID NO: 25.
TP-9 recombinant antigen fused with TP17-1 fragment, TP15 fragment, TP47-3 fragment and TP17-2 fragment from N terminal to C terminal, wherein TPThe connecting peptide between 17-1 and TP15 is (GGGGS)2, and the connecting peptide between TP15 and TP47-3 is GSG3The connecting peptide between SGSGSG, TP47-3 and TP17-2 is (AAY) 2; the nucleotide sequence of the prepared TP-9 recombinant antigen is shown in SEQ ID NO: 26, the amino acid sequence is shown in SEQ ID NO: 27.
example 2TP recombinant plasmid transformation and purification of recombinant protein induced expression
Transformation induced expression: the above-constructed expression plasmid was transformed into E.coli BL21 competent cells by heat shock method, plated on LB plates containing 200ug/ml of ampicillin sodium (AMP) (purchased from Solibao), and cultured at 37 ℃ for 16 hours. Selecting single colony, selecting positive bacterial strain which is identified correctly by PCR and double enzyme digestion of bacterial liquid, preserving the strain, inoculating the positive bacterial strain into LB culture medium containing 200ug/ml AMP, and performing shake culture at 37 ℃. To be OD600After reaching 0.6-0.8, adding 0.5mM IPTG (purchased from AMRESCO), carrying out induced culture at 28 ℃ for 4h, collecting bacteria, centrifuging at 6000rpm for 5min, and collecting bacteria precipitates; the pellet was resuspended in 50ml of 20mM PBS (pH 7.4), centrifuged at 6000rpm for 15min and the pellet collected.
Protein purification: resuspending the pellet in 20mM PBS (pH 7.4), performing low temperature pressure disruption, setting the pressure of a high pressure homogenizer at 850Bar, performing three cycles at 4 deg.C, centrifuging the disrupted product at 7500rpm for 30min, and collecting the supernatant. SDS-PAGE identifies recombinant protein expression. Purifying recombinant protein by nickel ion chelation of 6 HIS tag carried by PET32a vector, purifying recombinant protein by GST affinity chromatography column by GST tag carried by pGEX-4T-1 vector, purifying the collected primary pure protein by Q column, and purifying to obtain protein with purity of 95%. FIG. 1 shows a flow chart of the method for preparing TP recombinant antigen according to the present invention.
Example 3 preparation of a colloidal gold test strip by TP double antigen sandwich method
(1) Colloidal gold labeling
1mL of colloidal gold is put into a small glass cup, and a proper amount of 0.2M K is added into the small glass cup with stirring2CO3Adjusting the pH value to 7.6, and stirring for 1 min; adding 20 μ g of anti-TRX-tag monoclonal antibody (purchased from Shandong Shuojing biology), and stirring for 1 min; adding 100 μ L of 1% PEG20000, stirring for 1 min; centrifuging at 5000g for 10 min, discarding the supernatantThe precipitate was diluted with 100. mu.l of colloidal gold (20mM PB, 150mM NaCl, 0.1% PEG20000, 0.1% Triton X-100, 2% Sucrose, 0.01% NaN)3) Resuspending; finally, 5 μ g of PET32a-TP-Ag is added into 100 μ l of the colloidal gold labeled TRX monoclonal antibody compound, and the mixture is fully mixed and stored at 4 ℃. And (3) diluting the gold-labeled compound by 10 times of colloidal gold diluent, uniformly coating the diluted gold-labeled compound on glass fiber, and drying at 37 ℃ to prepare the gold-labeled pad.
(2) Nitrocellulose membrane (NC membrane) coating
The prepared TP recombinant antigen pGEX-4T-1-TP-1-Ag is diluted to a working concentration by a coating solution (20mM PBS 5% sucrose pH7.4) to be used as a coating raw material of a detection line (T line), and is lined on an NC membrane by using a membrane scribing instrument together with a coating raw material of a quality control line (C line) (goat anti-mouse IgG, purchased from Shandong Shuojing biology), and then is dried for 30min at 37 ℃, so that the solid phase of the raw material is completed.
(3) Assembly
And (3) assembling the solid-phase NC film with a sample pad, a gold label pad, a PVC bottom plate, absorbent paper and a Mark paste (the related sample pad, the gold label pad, the NC film, the PVC bottom plate, the absorbent paper and the Mark paste are purchased from Shandong Kanghua biology), and cutting into test strips with the thickness of 3-4 mm by using a slitter for later use. Fig. 2 shows a structure diagram of the colloidal gold test strip, and fig. 3 shows a result description of the colloidal gold test strip.
Example 4 application of colloidal gold test strip by TP double antigen sandwich method
435 cases of TP positive samples and 5000 cases of negative samples are detected by adopting the colloidal gold test strip prepared by the invention, and the control test strip is a treponema pallidum antibody detection test strip (purchased from Shandong Kanghua biology, registration number: national mechanical Standard 20153401404).
The detection results are shown in table 1, and from the detection results, the coating antigen is fixed: under the condition of pGEX-4T-1-TP-1, the sensitivity and the specificity of a test strip prepared by coupling and marking pET32a-TP-1 are respectively 99.7 percent and 99.78 percent; the sensitivity of the test strip prepared by using the coupling labeled antigen pET32a-TP-2 is 99.5 percent, and the specificity is 99.84 percent; the sensitivity of the test strip prepared by coupling labeled antigen pET32a-TP-3 is 99.3%, and the specificity is 99.80%; the sensitivity of the test strip prepared by using the coupling labeled antigen pET32a-TP-4 is 99.7%, and the specificity is 99.76%; the sensitivity of the test strip prepared by coupling labeled antigen pET32a-TP-5 is 100%, and the specificity is 99.68%; the sensitivity of the test strip prepared by using the coupling labeled antigen pET32a-TP-6 is 100%, and the specificity is 99.8%; the sensitivity of the test strip prepared by coupling labeled antigen pET32a-TP-7 is 100%, and the specificity is 99.82%; the sensitivity of the test strip prepared by using the coupling labeled antigen pET32a-TP-8 is 99.7%, and the specificity is 99.76%; the sensitivity of the test strip prepared by coupling labeled antigen pET32a-TP-9 is 99.5%, and the specificity is 99.74%.
As can be seen by the comparative analysis of the detection results, the self-made colloidal gold test strip has better specificity than the control test strip under the condition that the sensitivity of the self-made colloidal gold test strip is the same as that of the control test strip; or under the condition that the specificity is the same as that of the control test strip, the sensitivity of the test strip is superior to that of the control test strip; in most of the tests, the overall analysis of the sensitivity and specificity of the self-made colloidal gold test strip is superior to that of the control test strip.
TABLE 1 test results of self-made test strip and control test strip
Figure BDA0003409219800000081
Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art will understand that various changes, modifications and substitutions can be made without departing from the spirit and scope of the present invention. Any modification, equivalent replacement, or modification made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Sequence listing
<110> Qingdao Han Tang Biotech Co., Ltd
<120> TP recombinant antigen and preparation method and application thereof
<160> 27
<170> SIPOSequenceListing 1.0
<210> 1
<211> 134
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 1
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys
130
<210> 2
<211> 123
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 2
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met
115 120
<210> 3
<211> 121
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 3
Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe Asp
1 5 10 15
Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly Lys
20 25 30
Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe Lys
35 40 45
Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly Ile
50 55 60
Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu Lys
65 70 75 80
Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser Ser
85 90 95
Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg Arg
100 105 110
Gly Glu Lys Glu Ala Ile Ile Ser Arg
115 120
<210> 4
<211> 343
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 4
Met Phe Asp Ala Val Ser Arg Ala Thr His Gly His Gly Ala Phe Arg
1 5 10 15
Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly Glu Lys Val Leu Ser
20 25 30
Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys Lys Trp Glu Tyr Glu
35 40 45
Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala Ser Ala Ser Phe Gln
50 55 60
Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu Ala Val Glu Gly Ala
65 70 75 80
Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met Val Asp Ser Glu Glu
85 90 95
Tyr Lys Ile Thr Asn Val Lys Val His Gly Met Lys Phe Val Pro Val
100 105 110
Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys Glu Lys Phe His Phe
115 120 125
Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly Leu Lys Thr Met
130 135 140
Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val Ser Ser Met Glu Ser
145 150 155 160
Pro His Asp Leu Val Val Asp Thr Val Gly Thr Val Tyr His Ser Arg
165 170 175
Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys Arg Ala Asp Gly
180 185 190
Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val Met Asn Phe Asn
195 200 205
Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser Tyr Thr Asn Leu
210 215 220
Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser Trp Trp Lys Thr
225 230 235 240
Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr Gly Phe Asp Arg
245 250 255
Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr Leu Ile Ala Asn
260 265 270
Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu Pro Lys Tyr Glu
275 280 285
Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu Thr Ile Gly Gly
290 295 300
Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp Val Phe Ala Asp
305 310 315 320
Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser Leu Gly Gln Asn
325 330 335
Val Leu Ser Ala Asp Phe Thr
340
<210> 5
<211> 181
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 5
Val Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu
1 5 10 15
Lys Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr
20 25 30
Val Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala
35 40 45
Ser Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp
50 55 60
Ser Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn
65 70 75 80
Tyr Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu
85 90 95
Thr Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe
100 105 110
Leu Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val
115 120 125
Leu Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val
130 135 140
Asp Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val
145 150 155 160
Ser Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu
165 170 175
Tyr Thr Val Glu Val
180
<210> 6
<211> 414
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 6
Gly Ser Ser His His Glu Thr His Tyr Gly Tyr Ala Thr Leu Ser Tyr
1 5 10 15
Ala Asp Tyr Trp Ala Gly Glu Leu Gly Gln Ser Arg Asp Val Leu Leu
20 25 30
Ala Gly Asn Ala Glu Ala Asp Arg Ala Gly Asp Leu Asp Ala Gly Met
35 40 45
Phe Asp Ala Val Ser Arg Ala Thr His Gly His Gly Ala Phe Arg Gln
50 55 60
Gln Phe Gln Tyr Ala Val Glu Val Leu Gly Glu Lys Val Leu Ser Lys
65 70 75 80
Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys Lys Trp Glu Tyr Glu Thr
85 90 95
Asp Pro Ser Val Thr Lys Met Val Arg Ala Ser Ala Ser Phe Gln Asp
100 105 110
Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu Ala Val Glu Gly Ala Val
115 120 125
Ala Leu Ala Asp Arg Ala Ser Ser Phe Met Val Asp Ser Glu Glu Tyr
130 135 140
Lys Ile Thr Asn Val Lys Val His Gly Met Lys Phe Val Pro Val Ala
145 150 155 160
Val Pro His Glu Leu Lys Gly Ile Ala Lys Glu Lys Phe His Phe Val
165 170 175
Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly Leu Lys Thr Met Leu
180 185 190
Thr Glu Asp Ser Phe Ser Ala Arg Lys Val Ser Ser Met Glu Ser Pro
195 200 205
His Asp Leu Val Val Asp Thr Val Gly Thr Val Tyr His Ser Arg Phe
210 215 220
Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys Arg Ala Asp Gly Ser
225 230 235 240
Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val Met Asn Phe Asn Thr
245 250 255
Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser Tyr Thr Asn Leu Met
260 265 270
Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser Trp Trp Lys Thr Gly
275 280 285
Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr Gly Phe Asp Arg Phe
290 295 300
Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr Leu Ile Ala Asn Gly
305 310 315 320
Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu Pro Lys Tyr Glu Gly
325 330 335
Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu Thr Ile Gly Gly Ala
340 345 350
Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp Val Phe Ala Asp Gly
355 360 365
Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser Leu Gly Gln Asn Val
370 375 380
Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu Tyr Thr Val Glu Val Arg
385 390 395 400
Phe Lys Glu Phe Gly Ser Val Arg Ala Lys Val Val Ala Gln
405 410
<210> 7
<211> 402
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 7
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aa 402
<210> 8
<211> 366
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 8
tcctcttcta tcccgaacgg tacctaccgt gctacctacc aggacttcga cgaaaacggt 60
tggaaagact tcctggaagt taccttcgac ggtggtaaaa tggttcaggt tgtttacgac 120
taccagcaca aagaaggtcg tttcaaatct caggacgctg actaccaccg tgttatgtac 180
gcttcttctg gtatcggtcc ggagaaggcg ttccgtgaac tggcggacgc tctgctggaa 240
aaaggtaacc cggaaatggt tgacgttgtt acaggtgcga ccgtcagctc tcagtctttc 300
cgtcgtctgg gtgctgctct gctgcagtct gctcgtcgtg gtgaaaaaga agctatcatc 360
tctcgt 366
<210> 9
<211> 1242
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 9
ggttctagtc atcacgaaac ccattacggt tacgcgaccc tgagttacgc agattattgg 60
gcaggcgaac tgggtcaaag tcgcgacgtt ctgctggcag gtaacgcaga agcagatcgc 120
gcaggtgatc tggacgcagg tatgtttgac gcagtttctc gtgcaaccca cggtcacggc 180
gcatttcgtc aacagtttca gtacgcggtg gaagttctgg gcgaaaaagt gctgagcaaa 240
caggaaaccg aagatagccg cggtcgcaaa aaatgggaat acgaaaccga cccgagcgtt 300
accaaaatgg ttcgcgcaag cgcaagcttt caagatctgg gcgaagacgg cgaaatcaaa 360
ttcgaagcag tggaaggcgc agttgcactg gcagatcgcg caagtagctt tatggtggat 420
agcgaagagt acaaaatcac caacgtgaaa gtgcacggca tgaaatttgt tccggttgca 480
gttccgcacg aactgaaagg catcgcgaaa gagaaattcc attttgtgga agatagccgc 540
gtgaccgaaa acaccaacgg cctgaaaacc atgctgaccg aagatagctt tagcgcgcgt 600
aaagtgagca gtatggaaag tccgcacgat ctggttgttg ataccgtagg taccgtttac 660
cactctcgtt tcggttctga cgctgaggct tctgttatgc tgaaacgtgc tgacggttct 720
gaactgtctc accgtgagtt catcgactac gttatgaact tcaacaccgt tcgttacgac 780
tactacggtg acgacgcttc ttacaccaac ctgatggctt cttacggtac caaacactct 840
gctgactctt ggtggaaaac cggtcgtgtt ccgcgtatct cttgcggtat caactacggt 900
ttcgaccgtt tcaaaggttc tggtccgggt tactaccgtc tgaccctgat cgctaacggt 960
taccgtgacg ttgttgctga cgttcgtttc ctgccgaaat acgaaggtaa catcgacatc 1020
ggtctgaaag gtaaagttct gaccatcggt ggtgctgacg ctgaaaccct gatggacgct 1080
gctgttgacg ttttcgctga cggtcagccg aaactggttt ctgaccaggc tgtttctctg 1140
ggtcagaacg ttctgtctgc tgacttcacc ccgggtaccg aatacaccgt tgaagttcgt 1200
ttcaaagaat ttggttctgt tcgtgctaaa gttgttgctc ag 1242
<210> 10
<211> 2075
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 10
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 420
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 480
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 540
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 600
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 660
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 720
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 780
gaagctatca tctctcgtgg tctggtggtg gttctggttc tggttctggt ggttctagtc 840
atcacgaaac ccattacggt tacgcgaccc tgagttacgc agattattgg gcaggcgaac 900
tgggtcaaag tcgcgacgtt ctgctggcag gtaacgcaga agcagatcgc gcaggtgatc 960
tggacgcagg tatgtttgac gcagtttctc gtgcaaccca cggtcacggc gcatttcgtc 1020
aacagtttca gtacgcggtg gaagttctgg gcgaaaaagt gctgagcaaa caggaaaccg 1080
aagatagccg cggtcgcaaa aaatgggaat acgaaaccga cccgagcgtt accaaaatgg 1140
ttcgcgcaag cgcaagcttt caagatctgg gcgaagacgg cgaaatcaaa ttcgaagcag 1200
tggaaggcgc agttgcactg gcagatcgcg caagtagctt tatggtggat agcgaagagt 1260
acaaaatcac caacgtgaaa gtgcacggca tgaaatttgt tccggttgca gttccgcacg 1320
aactgaaagg catcgcgaaa gagaaattcc attttgtgga agatagccgc gtgaccgaaa 1380
acaccaacgg cctgaaaacc atgctgaccg aagatagctt tagcgcgcgt aaagtgagca 1440
gtatggaaag tccgcacgat ctggttgttg ataccgtagg taccgtttac cactctcgtt 1500
tcggttctga cgctgaggct tctgttatgc tgaaacgtgc tgacggttct gaactgtctc 1560
accgtgagtt catcgactac gttatgaact tcaacaccgt tcgttacgac tactacggtg 1620
acgacgcttc ttacaccaac ctgatggctt cttacggtac caaacactct gctgactctt 1680
ggtggaaaac cggtcgtgtt ccgcgtatct cttgcggtat caactacggt ttcgaccgtt 1740
tcaaaggttc tggtccgggt tactaccgtc tgaccctgat cgctaacggt taccgtgacg 1800
ttgttgctga cgttcgtttc ctgccgaaat acgaaggtaa catcgacatc ggtctgaaag 1860
gtaaagttct gaccatcggt ggtgctgacg ctgaaaccct gatggacgct gctgttgacg 1920
ttttcgctga cggtcagccg aaactggttt ctgaccaggc tgtttctctg ggtcagaacg 1980
ttctgtctgc tgacttcacc ccgggtaccg aatacaccgt tgaagttcgt ttcaaagaat 2040
ttggttctgt tcgtgctaaa gttgttgctc agtaa 2075
<210> 11
<211> 691
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 11
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
130 135 140
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
145 150 155 160
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
165 170 175
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
180 185 190
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
195 200 205
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
210 215 220
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
225 230 235 240
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
245 250 255
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser
260 265 270
Gly Ser Gly Ser Gly Gly Ser Ser His His Glu Thr His Tyr Gly Tyr
275 280 285
Ala Thr Leu Ser Tyr Ala Asp Tyr Trp Ala Gly Glu Leu Gly Gln Ser
290 295 300
Arg Asp Val Leu Leu Ala Gly Asn Ala Glu Ala Asp Arg Ala Gly Asp
305 310 315 320
Leu Asp Ala Gly Met Phe Asp Ala Val Ser Arg Ala Thr His Gly His
325 330 335
Gly Ala Phe Arg Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly Glu
340 345 350
Lys Val Leu Ser Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys Lys
355 360 365
Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala Ser
370 375 380
Ala Ser Phe Gln Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu Ala
385 390 395 400
Val Glu Gly Ala Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met Val
405 410 415
Asp Ser Glu Glu Tyr Lys Ile Thr Asn Val Lys Val His Gly Met Lys
420 425 430
Phe Val Pro Val Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys Glu
435 440 445
Lys Phe His Phe Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly
450 455 460
Leu Lys Thr Met Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val Ser
465 470 475 480
Ser Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr Val
485 490 495
Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys
500 505 510
Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val
515 520 525
Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser
530 535 540
Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser
545 550 555 560
Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr
565 570 575
Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr
580 585 590
Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu
595 600 605
Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu
610 615 620
Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp
625 630 635 640
Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser
645 650 655
Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu Tyr
660 665 670
Thr Val Glu Val Arg Phe Lys Glu Phe Gly Ser Val Arg Ala Lys Val
675 680 685
Val Ala Gln
690
<210> 12
<211> 1371
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 12
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 420
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 480
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 540
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 600
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 660
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 720
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 780
gaagctatca tctctcgtgg ttctggtggt ggttctggtt ctggtggtgt ttaccactct 840
cgtttcggtt ctgacgctga ggcttctgtt atgctgaaac gtgctgacgg ttctgaactg 900
tctcaccgtg agttcatcga ctacgttatg aacttcaaca ccgttcgtta cgactactac 960
ggtgacgacg cttcttacac caacctgatg gcttcttacg gtaccaaaca ctctgctgac 1020
tcttggtgga aaaccggtcg tgttccgcgt atctcttgcg gtatcaacta cggtttcgac 1080
cgtttcaaag gttctggtcc gggttactac cgtctgaccc tgatcgctaa cggttaccgt 1140
gacgttgttg ctgacgttcg tttcctgccg aaatacgaag gtaacatcga catcggtctg 1200
aaaggtaaag ttctgaccat cggtggtgct gacgctgaaa ccctgatgga cgctgctgtt 1260
gacgttttcg ctgacggtca gccgaaactg gtttctgacc aggctgtttc tctgggtcag 1320
aacgttctgt ctgctgactt caccccgggt accgaataca ccgttgaagt t 1371
<210> 13
<211> 459
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 13
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
130 135 140
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
145 150 155 160
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
165 170 175
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
180 185 190
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
195 200 205
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
210 215 220
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
225 230 235 240
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
245 250 255
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser
260 265 270
Gly Ser Gly Ser Gly Gly Val Tyr His Ser Arg Phe Gly Ser Asp Ala
275 280 285
Glu Ala Ser Val Met Leu Lys Arg Ala Asp Gly Ser Glu Leu Ser His
290 295 300
Arg Glu Phe Ile Asp Tyr Val Met Asn Phe Asn Thr Val Arg Tyr Asp
305 310 315 320
Tyr Tyr Gly Asp Asp Ala Ser Tyr Thr Asn Leu Met Ala Ser Tyr Gly
325 330 335
Thr Lys His Ser Ala Asp Ser Trp Trp Lys Thr Gly Arg Val Pro Arg
340 345 350
Ile Ser Cys Gly Ile Asn Tyr Gly Phe Asp Arg Phe Lys Gly Ser Gly
355 360 365
Pro Gly Tyr Tyr Arg Leu Thr Leu Ile Ala Asn Gly Tyr Arg Asp Val
370 375 380
Val Ala Asp Val Arg Phe Leu Pro Lys Tyr Glu Gly Asn Ile Asp Ile
385 390 395 400
Gly Leu Lys Gly Lys Val Leu Thr Ile Gly Gly Ala Asp Ala Glu Thr
405 410 415
Leu Met Asp Ala Ala Val Asp Val Phe Ala Asp Gly Gln Pro Lys Leu
420 425 430
Val Ser Asp Gln Ala Val Ser Leu Gly Gln Asn Val Leu Ser Ala Asp
435 440 445
Phe Thr Pro Gly Thr Glu Tyr Thr Val Glu Val
450 455
<210> 14
<211> 1860
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 14
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 420
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 480
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 540
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 600
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 660
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 720
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 780
gaagctatca tctctcgtgg ttctggtggt ggttctggtt ctggttctgg tatgtttgac 840
gcagtttctc gtgcaaccca cggtcacggc gcatttcgtc aacagtttca gtacgcggtg 900
gaagttctgg gcgaaaaagt gctgagcaaa caggaaaccg aagatagccg cggtcgcaaa 960
aaatgggaat acgaaaccga cccgagcgtt accaaaatgg ttcgcgcaag cgcaagcttt 1020
caagatctgg gcgaagacgg cgaaatcaaa ttcgaagcag tggaaggcgc agttgcactg 1080
gcagatcgcg caagtagctt tatggtggat agcgaagagt acaaaatcac caacgtgaaa 1140
gtgcacggca tgaaatttgt tccggttgca gttccgcacg aactgaaagg catcgcgaaa 1200
gagaaattcc attttgtgga agatagccgc gtgaccgaaa acaccaacgg cctgaaaacc 1260
atgctgaccg aagatagctt tagcgcgcgt aaagtgagca gtatggaaag tccgcacgat 1320
ctggttgttg ataccgtagg taccgtttac cactctcgtt tcggttctga cgctgaggct 1380
tctgttatgc tgaaacgtgc tgacggttct gaactgtctc accgtgagtt catcgactac 1440
gttatgaact tcaacaccgt tcgttacgac tactacggtg acgacgcttc ttacaccaac 1500
ctgatggctt cttacggtac caaacactct gctgactctt ggtggaaaac cggtcgtgtt 1560
ccgcgtatct cttgcggtat caactacggt ttcgaccgtt tcaaaggttc tggtccgggt 1620
tactaccgtc tgaccctgat cgctaacggt taccgtgacg ttgttgctga cgttcgtttc 1680
ctgccgaaat acgaaggtaa catcgacatc ggtctgaaag gtaaagttct gaccatcggt 1740
ggtgctgacg ctgaaaccct gatggacgct gctgttgacg ttttcgctga cggtcagccg 1800
aaactggttt ctgaccaggc tgtttctctg ggtcagaacg ttctgtctgc tgacttcacc 1860
<210> 15
<211> 620
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 15
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
130 135 140
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
145 150 155 160
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
165 170 175
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
180 185 190
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
195 200 205
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
210 215 220
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
225 230 235 240
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
245 250 255
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser
260 265 270
Gly Ser Gly Ser Gly Met Phe Asp Ala Val Ser Arg Ala Thr His Gly
275 280 285
His Gly Ala Phe Arg Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly
290 295 300
Glu Lys Val Leu Ser Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys
305 310 315 320
Lys Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala
325 330 335
Ser Ala Ser Phe Gln Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu
340 345 350
Ala Val Glu Gly Ala Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met
355 360 365
Val Asp Ser Glu Glu Tyr Lys Ile Thr Asn Val Lys Val His Gly Met
370 375 380
Lys Phe Val Pro Val Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys
385 390 395 400
Glu Lys Phe His Phe Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn
405 410 415
Gly Leu Lys Thr Met Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val
420 425 430
Ser Ser Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr
435 440 445
Val Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu
450 455 460
Lys Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr
465 470 475 480
Val Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala
485 490 495
Ser Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp
500 505 510
Ser Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn
515 520 525
Tyr Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu
530 535 540
Thr Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe
545 550 555 560
Leu Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val
565 570 575
Leu Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val
580 585 590
Asp Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val
595 600 605
Ser Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr
610 615 620
<210> 16
<211> 1374
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 16
tcctcttcta tcccgaacgg tacctaccgt gctacctacc aggacttcga cgaaaacggt 60
tggaaagact tcctggaagt taccttcgac ggtggtaaaa tggttcaggt tgtttacgac 120
taccagcaca aagaaggtcg tttcaaatct caggacgctg actaccaccg tgttatgtac 180
gcttcttctg gtatcggtcc ggagaaggcg ttccgtgaac tggcggacgc tctgctggaa 240
aaaggtaacc cggaaatggt tgacgttgtt acaggtgcga ccgtcagctc tcagtctttc 300
cgtcgtctgg gtgctgctct gctgcagtct gctcgtcgtg gtgaaaaaga agctatcatc 360
tctcgtgctg cttacgctgc ttacgtttct tgcaccaccg tttgcccgca cgctggtaaa 420
gctaaagctg aaaaagttga agctgctctg aaaggtggta tcttccgtgg taccctgccg 480
gctgctgacg ctccgggtat cgacaccacc gttaccttca acgctgacgg taccgctcag 540
aaagttgaac tggctctgga aaaaaaatct gctccgtctc cgctgaccta ccgtggtacc 600
tggatggttc gtgaagacgg tatcgttgaa ctgtctctgg tttcttctga acagtctaaa 660
gctccgcacg aaaaagaact gtacgaactg atcgactcta actctgttcg ttacatgggt 720
gctccgggtg ctggtaaacc gtctaaagaa atggctccgt tctacgttct gaaaaaaacc 780
aaaaaaggtg gtggtggttc tggtggtggt ggttctggtg gtggtggttc tgtttaccac 840
tctcgtttcg gttctgacgc tgaggcttct gttatgctga aacgtgctga cggttctgaa 900
ctgtctcacc gtgagttcat cgactacgtt atgaacttca acaccgttcg ttacgactac 960
tacggtgacg acgcttctta caccaacctg atggcttctt acggtaccaa acactctgct 1020
gactcttggt ggaaaaccgg tcgtgttccg cgtatctctt gcggtatcaa ctacggtttc 1080
gaccgtttca aaggttctgg tccgggttac taccgtctga ccctgatcgc taacggttac 1140
cgtgacgttg ttgctgacgt tcgtttcctg ccgaaatacg aaggtaacat cgacatcggt 1200
ctgaaaggta aagttctgac catcggtggt gctgacgctg aaaccctgat ggacgctgct 1260
gttgacgttt tcgctgacgg tcagccgaaa ctggtttctg accaggctgt ttctctgggt 1320
cagaacgttc tgtctgctga cttcaccccg ggtaccgaat acaccgttga agtt 1374
<210> 17
<211> 333
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 17
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
1 5 10 15
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
20 25 30
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
35 40 45
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
50 55 60
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
65 70 75 80
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
85 90 95
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
100 105 110
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Ala Ala Tyr Ala Ala Tyr
115 120 125
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
130 135 140
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
145 150 155 160
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
165 170 175
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
180 185 190
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
195 200 205
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
210 215 220
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
225 230 235 240
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
245 250 255
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
260 265 270
Gly Gly Gly Gly Ser Val Tyr His Ser Arg Phe Gly Ser Asp Ala Glu
275 280 285
Ala Ser Val Met Leu Lys Arg Ala Asp Gly Ser Glu Leu Ser His Arg
290 295 300
Glu Phe Ile Asp Tyr Val Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr
305 310 315 320
Tyr Gly Asp Asp Ala Ser Tyr Thr Asn Leu Met Ala Ser
325 330
<210> 18
<211> 1860
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 18
tcctcttcta tcccgaacgg tacctaccgt gctacctacc aggacttcga cgaaaacggt 60
tggaaagact tcctggaagt taccttcgac ggtggtaaaa tggttcaggt tgtttacgac 120
taccagcaca aagaaggtcg tttcaaatct caggacgctg actaccaccg tgttatgtac 180
gcttcttctg gtatcggtcc ggagaaggcg ttccgtgaac tggcggacgc tctgctggaa 240
aaaggtaacc cggaaatggt tgacgttgtt acaggtgcga ccgtcagctc tcagtctttc 300
cgtcgtctgg gtgctgctct gctgcagtct gctcgtcgtg gtgaaaaaga agctatcatc 360
tctcgtgctg cttacgctgc ttacgtttct tgcaccaccg tttgcccgca cgctggtaaa 420
gctaaagctg aaaaagttga agctgctctg aaaggtggta tcttccgtgg taccctgccg 480
gctgctgacg ctccgggtat cgacaccacc gttaccttca acgctgacgg taccgctcag 540
aaagttgaac tggctctgga aaaaaaatct gctccgtctc cgctgaccta ccgtggtacc 600
tggatggttc gtgaagacgg tatcgttgaa ctgtctctgg tttcttctga acagtctaaa 660
gctccgcacg aaaaagaact gtacgaactg atcgactcta actctgttcg ttacatgggt 720
gctccgggtg ctggtaaacc gtctaaagaa atggctccgt tctacgttct gaaaaaaacc 780
aaaaaaggtg gtggtggttc tggtggtggt ggttctggtg gtggtggttc tatgtttgac 840
gcagtttctc gtgcaaccca cggtcacggc gcatttcgtc aacagtttca gtacgcggtg 900
gaagttctgg gcgaaaaagt gctgagcaaa caggaaaccg aagatagccg cggtcgcaaa 960
aaatgggaat acgaaaccga cccgagcgtt accaaaatgg ttcgcgcaag cgcaagcttt 1020
caagatctgg gcgaagacgg cgaaatcaaa ttcgaagcag tggaaggcgc agttgcactg 1080
gcagatcgcg caagtagctt tatggtggat agcgaagagt acaaaatcac caacgtgaaa 1140
gtgcacggca tgaaatttgt tccggttgca gttccgcacg aactgaaagg catcgcgaaa 1200
gagaaattcc attttgtgga agatagccgc gtgaccgaaa acaccaacgg cctgaaaacc 1260
atgctgaccg aagatagctt tagcgcgcgt aaagtgagca gtatggaaag tccgcacgat 1320
ctggttgttg ataccgtagg taccgtttac cactctcgtt tcggttctga cgctgaggct 1380
tctgttatgc tgaaacgtgc tgacggttct gaactgtctc accgtgagtt catcgactac 1440
gttatgaact tcaacaccgt tcgttacgac tactacggtg acgacgcttc ttacaccaac 1500
ctgatggctt cttacggtac caaacactct gctgactctt ggtggaaaac cggtcgtgtt 1560
ccgcgtatct cttgcggtat caactacggt ttcgaccgtt tcaaaggttc tggtccgggt 1620
tactaccgtc tgaccctgat cgctaacggt taccgtgacg ttgttgctga cgttcgtttc 1680
ctgccgaaat acgaaggtaa catcgacatc ggtctgaaag gtaaagttct gaccatcggt 1740
ggtgctgacg ctgaaaccct gatggacgct gctgttgacg ttttcgctga cggtcagccg 1800
aaactggttt ctgaccaggc tgtttctctg ggtcagaacg ttctgtctgc tgacttcacc 1860
<210> 19
<211> 620
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 19
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
1 5 10 15
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
20 25 30
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
35 40 45
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
50 55 60
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
65 70 75 80
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
85 90 95
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
100 105 110
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Ala Ala Tyr Ala Ala Tyr
115 120 125
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
130 135 140
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
145 150 155 160
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
165 170 175
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
180 185 190
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
195 200 205
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
210 215 220
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
225 230 235 240
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
245 250 255
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
260 265 270
Gly Gly Gly Gly Ser Met Phe Asp Ala Val Ser Arg Ala Thr His Gly
275 280 285
His Gly Ala Phe Arg Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly
290 295 300
Glu Lys Val Leu Ser Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys
305 310 315 320
Lys Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala
325 330 335
Ser Ala Ser Phe Gln Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu
340 345 350
Ala Val Glu Gly Ala Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met
355 360 365
Val Asp Ser Glu Glu Tyr Lys Ile Thr Asn Val Lys Val His Gly Met
370 375 380
Lys Phe Val Pro Val Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys
385 390 395 400
Glu Lys Phe His Phe Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn
405 410 415
Gly Leu Lys Thr Met Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val
420 425 430
Ser Ser Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr
435 440 445
Val Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu
450 455 460
Lys Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr
465 470 475 480
Val Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala
485 490 495
Ser Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp
500 505 510
Ser Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn
515 520 525
Tyr Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu
530 535 540
Thr Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe
545 550 555 560
Leu Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val
565 570 575
Leu Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val
580 585 590
Asp Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val
595 600 605
Ser Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr
610 615 620
<210> 20
<211> 1629
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 20
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aagctgctta cgctgcttac 420
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 480
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 540
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 600
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtggt 660
ggtggttctg gtggtggtgg ttcttcctct tctatcccga acggtaccta ccgtgctacc 720
taccaggact tcgacgaaaa cggttggaaa gacttcctgg aagttacctt cgacggtggt 780
aaaatggttc aggttgttta cgactaccag cacaaagaag gtcgtttcaa atctcaggac 840
gctgactacc accgtgttat gtacgcttct tctggtatcg gtccggagaa ggcgttccgt 900
gaactggcgg acgctctgct ggaaaaaggt aacccggaaa tggttgacgt tgttacaggt 960
gcgaccgtca gctctcagtc tttccgtcgt ctgggtgctg ctctgctgca gtctgctcgt 1020
cgtggtgaaa aagaagctat catctctcgt ggttctggtg gtggttctgg ttctggttct 1080
ggtggtgttt accactctcg tttcggttct gacgctgagg cttctgttat gctgaaacgt 1140
gctgacggtt ctgaactgtc tcaccgtgag ttcatcgact acgttatgaa cttcaacacc 1200
gttcgttacg actactacgg tgacgacgct tcttacacca acctgatggc ttcttacggt 1260
accaaacact ctgctgactc ttggtggaaa accggtcgtg ttccgcgtat ctcttgcggt 1320
atcaactacg gtttcgaccg tttcaaaggt tctggtccgg gttactaccg tctgaccctg 1380
atcgctaacg gttaccgtga cgttgttgct gacgttcgtt tcctgccgaa atacgaaggt 1440
aacatcgaca tcggtctgaa aggtaaagtt ctgaccatcg gtggtgctga cgctgaaacc 1500
ctgatggacg ctgctgttga cgttttcgct gacggtcagc cgaaactggt ttctgaccag 1560
gctgtttctc tgggtcagaa cgttctgtct gctgacttca ccccgggtac cgaatacacc 1620
gttgaagtt 1629
<210> 21
<211> 543
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 21
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Ala Ala Tyr Ala Ala Tyr Val Ser Cys Thr
130 135 140
Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu Lys Val Glu Ala
145 150 155 160
Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro Ala Ala Asp Ala
165 170 175
Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp Gly Thr Ala Gln
180 185 190
Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro Ser Pro Leu Thr
195 200 205
Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Gly Gly Gly Ser Gly
210 215 220
Gly Gly Gly Ser Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr
225 230 235 240
Tyr Gln Asp Phe Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr
245 250 255
Phe Asp Gly Gly Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys
260 265 270
Glu Gly Arg Phe Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr
275 280 285
Ala Ser Ser Gly Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp
290 295 300
Ala Leu Leu Glu Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly
305 310 315 320
Ala Thr Val Ser Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu
325 330 335
Gln Ser Ala Arg Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser
340 345 350
Gly Gly Gly Ser Gly Ser Gly Ser Gly Gly Val Tyr His Ser Arg Phe
355 360 365
Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys Arg Ala Asp Gly Ser
370 375 380
Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val Met Asn Phe Asn Thr
385 390 395 400
Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser Tyr Thr Asn Leu Met
405 410 415
Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser Trp Trp Lys Thr Gly
420 425 430
Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr Gly Phe Asp Arg Phe
435 440 445
Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr Leu Ile Ala Asn Gly
450 455 460
Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu Pro Lys Tyr Glu Gly
465 470 475 480
Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu Thr Ile Gly Gly Ala
485 490 495
Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp Val Phe Ala Asp Gly
500 505 510
Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser Leu Gly Gln Asn Val
515 520 525
Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu Tyr Thr Val Glu Val
530 535 540
<210> 22
<211> 1797
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 22
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aagctgctta cgctgcttac 420
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 480
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 540
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 600
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 660
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 720
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 780
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 840
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 900
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 960
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 1020
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 1080
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 1140
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 1200
gaagctatca tctctcgtgg ttctggtggt ggttctggtt ctggttctgg tggtgtttac 1260
cactctcgtt tcggttctga cgctgaggct tctgttatgc tgaaacgtgc tgacggttct 1320
gaactgtctc accgtgagtt catcgactac gttatgaact tcaacaccgt tcgttacgac 1380
tactacggtg acgacgcttc ttacaccaac ctgatggctt cttacggtac caaacactct 1440
gctgactctt ggtggaaaac cggtcgtgtt ccgcgtatct cttgcggtat caactacggt 1500
ttcgaccgtt tcaaaggttc tggtccgggt tactaccgtc tgaccctgat cgctaacggt 1560
taccgtgacg ttgttgctga cgttcgtttc ctgccgaaat acgaaggtaa catcgacatc 1620
ggtctgaaag gtaaagttct gaccatcggt ggtgctgacg ctgaaaccct gatggacgct 1680
gctgttgacg ttttcgctga cggtcagccg aaactggttt ctgaccaggc tgtttctctg 1740
ggtcagaacg ttctgtctgc tgacttcacc ccgggtaccg aatacaccgt tgaagtt 1797
<210> 23
<211> 599
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 23
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Ala Ala Tyr Ala Ala Tyr Val Ser Cys Thr
130 135 140
Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu Lys Val Glu Ala
145 150 155 160
Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro Ala Ala Asp Ala
165 170 175
Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp Gly Thr Ala Gln
180 185 190
Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro Ser Pro Leu Thr
195 200 205
Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile Val Glu Leu Ser
210 215 220
Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu Lys Glu Leu Tyr
225 230 235 240
Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly Ala Pro Gly Ala
245 250 255
Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val Leu Lys Lys Thr
260 265 270
Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Ser Ser Ser Ile
275 280 285
Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe Asp Glu Asn Gly
290 295 300
Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly Lys Met Val Gln
305 310 315 320
Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe Lys Ser Gln Asp
325 330 335
Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly Ile Gly Pro Glu
340 345 350
Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu Lys Gly Asn Pro
355 360 365
Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser Ser Gln Ser Phe
370 375 380
Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg Arg Gly Glu Lys
385 390 395 400
Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser Gly Ser Gly Ser
405 410 415
Gly Gly Val Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val
420 425 430
Met Leu Lys Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile
435 440 445
Asp Tyr Val Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp
450 455 460
Asp Ala Ser Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser
465 470 475 480
Ala Asp Ser Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly
485 490 495
Ile Asn Tyr Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr
500 505 510
Arg Leu Thr Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val
515 520 525
Arg Phe Leu Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly
530 535 540
Lys Val Leu Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala
545 550 555 560
Ala Val Asp Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln
565 570 575
Ala Val Ser Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr Pro Gly
580 585 590
Thr Glu Tyr Thr Val Glu Val
595
<210> 24
<211> 2493
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 24
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 420
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 480
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 540
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 600
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 660
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 720
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 780
gaagctatca tctctcgtgg ttctggtggt ggttctggtt ctggttctgg tggttctagt 840
catcacgaaa cccattacgg ttacgcgacc ctgagttacg cagattattg ggcaggcgaa 900
ctgggtcaaa gtcgcgacgt tctgctggca ggtaacgcag aagcagatcg cgcaggtgat 960
ctggacgcag gtatgtttga cgcagtttct cgtgcaaccc acggtcacgg cgcatttcgt 1020
caacagtttc agtacgcggt ggaagttctg ggcgaaaaag tgctgagcaa acaggaaacc 1080
gaagatagcc gcggtcgcaa aaaatgggaa tacgaaaccg acccgagcgt taccaaaatg 1140
gttcgcgcaa gcgcaagctt tcaagatctg ggcgaagacg gcgaaatcaa attcgaagca 1200
gtggaaggcg cagttgcact ggcagatcgc gcaagtagct ttatggtgga tagcgaagag 1260
tacaaaatca ccaacgtgaa agtgcacggc atgaaatttg ttccggttgc agttccgcac 1320
gaactgaaag gcatcgcgaa agagaaattc cattttgtgg aagatagccg cgtgaccgaa 1380
aacaccaacg gcctgaaaac catgctgacc gaagatagct ttagcgcgcg taaagtgagc 1440
agtatggaaa gtccgcacga tctggttgtt gataccgtag gtaccgttta ccactctcgt 1500
ttcggttctg acgctgaggc ttctgttatg ctgaaacgtg ctgacggttc tgaactgtct 1560
caccgtgagt tcatcgacta cgttatgaac ttcaacaccg ttcgttacga ctactacggt 1620
gacgacgctt cttacaccaa cctgatggct tcttacggta ccaaacactc tgctgactct 1680
tggtggaaaa ccggtcgtgt tccgcgtatc tcttgcggta tcaactacgg tttcgaccgt 1740
ttcaaaggtt ctggtccggg ttactaccgt ctgaccctga tcgctaacgg ttaccgtgac 1800
gttgttgctg acgttcgttt cctgccgaaa tacgaaggta acatcgacat cggtctgaaa 1860
ggtaaagttc tgaccatcgg tggtgctgac gctgaaaccc tgatggacgc tgctgttgac 1920
gttttcgctg acggtcagcc gaaactggtt tctgaccagg ctgtttctct gggtcagaac 1980
gttctgtctg ctgacttcac cccgggtacc gaatacaccg ttgaagttcg tttcaaagaa 2040
tttggttctg ttcgtgctaa agttgttgct caggctgctt acgctgctta cgtttcttgc 2100
accaccgttt gcccgcacgc tggtaaagct aaagctgaaa aagttgaagc tgctctgaaa 2160
ggtggtatct tccgtggtac cctgccggct gctgacgctc cgggtatcga caccaccgtt 2220
accttcaacg ctgacggtac cgctcagaaa gttgaactgg ctctggaaaa aaaatctgct 2280
ccgtctccgc tgacctaccg tggtacctgg atggttcgtg aagacggtat cgttgaactg 2340
tctctggttt cttctgaaca gtctaaagct ccgcacgaaa aagaactgta cgaactgatc 2400
gactctaact ctgttcgtta catgggtgct ccgggtgctg gtaaaccgtc taaagaaatg 2460
gctccgttct acgttctgaa aaaaaccaaa aaa 2493
<210> 25
<211> 831
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 25
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
130 135 140
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
145 150 155 160
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
165 170 175
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
180 185 190
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
195 200 205
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
210 215 220
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
225 230 235 240
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
245 250 255
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser
260 265 270
Gly Ser Gly Ser Gly Gly Ser Ser His His Glu Thr His Tyr Gly Tyr
275 280 285
Ala Thr Leu Ser Tyr Ala Asp Tyr Trp Ala Gly Glu Leu Gly Gln Ser
290 295 300
Arg Asp Val Leu Leu Ala Gly Asn Ala Glu Ala Asp Arg Ala Gly Asp
305 310 315 320
Leu Asp Ala Gly Met Phe Asp Ala Val Ser Arg Ala Thr His Gly His
325 330 335
Gly Ala Phe Arg Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly Glu
340 345 350
Lys Val Leu Ser Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys Lys
355 360 365
Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala Ser
370 375 380
Ala Ser Phe Gln Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu Ala
385 390 395 400
Val Glu Gly Ala Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met Val
405 410 415
Asp Ser Glu Glu Tyr Lys Ile Thr Asn Val Lys Val His Gly Met Lys
420 425 430
Phe Val Pro Val Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys Glu
435 440 445
Lys Phe His Phe Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly
450 455 460
Leu Lys Thr Met Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val Ser
465 470 475 480
Ser Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr Val
485 490 495
Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys
500 505 510
Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val
515 520 525
Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser
530 535 540
Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser
545 550 555 560
Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr
565 570 575
Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr
580 585 590
Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu
595 600 605
Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu
610 615 620
Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp
625 630 635 640
Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser
645 650 655
Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu Tyr
660 665 670
Thr Val Glu Val Arg Phe Lys Glu Phe Gly Ser Val Arg Ala Lys Val
675 680 685
Val Ala Gln Ala Ala Tyr Ala Ala Tyr Val Ser Cys Thr Thr Val Cys
690 695 700
Pro His Ala Gly Lys Ala Lys Ala Glu Lys Val Glu Ala Ala Leu Lys
705 710 715 720
Gly Gly Ile Phe Arg Gly Thr Leu Pro Ala Ala Asp Ala Pro Gly Ile
725 730 735
Asp Thr Thr Val Thr Phe Asn Ala Asp Gly Thr Ala Gln Lys Val Glu
740 745 750
Leu Ala Leu Glu Lys Lys Ser Ala Pro Ser Pro Leu Thr Tyr Arg Gly
755 760 765
Thr Trp Met Val Arg Glu Asp Gly Ile Val Glu Leu Ser Leu Val Ser
770 775 780
Ser Glu Gln Ser Lys Ala Pro His Glu Lys Glu Leu Tyr Glu Leu Ile
785 790 795 800
Asp Ser Asn Ser Val Arg Tyr Met Gly Ala Pro Gly Ala Gly Lys Pro
805 810 815
Ser Lys Glu Met Ala Pro Phe Tyr Val Leu Lys Lys Thr Lys Lys
820 825 830
<210> 26
<211> 2325
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 26
gtttcttgca ccaccgtttg cccgcacgct ggtaaagcta aagctgaaaa agttgaagct 60
gctctgaaag gtggtatctt ccgtggtacc ctgccggctg ctgacgctcc gggtatcgac 120
accaccgtta ccttcaacgc tgacggtacc gctcagaaag ttgaactggc tctggaaaaa 180
aaatctgctc cgtctccgct gacctaccgt ggtacctgga tggttcgtga agacggtatc 240
gttgaactgt ctctggtttc ttctgaacag tctaaagctc cgcacgaaaa agaactgtac 300
gaactgatcg actctaactc tgttcgttac atgggtgctc cgggtgctgg taaaccgtct 360
aaagaaatgg ctccgttcta cgttctgaaa aaaaccaaaa aaggtggtgg tggttctggt 420
ggtggtggtt cttcctcttc tatcccgaac ggtacctacc gtgctaccta ccaggacttc 480
gacgaaaacg gttggaaaga cttcctggaa gttaccttcg acggtggtaa aatggttcag 540
gttgtttacg actaccagca caaagaaggt cgtttcaaat ctcaggacgc tgactaccac 600
cgtgttatgt acgcttcttc tggtatcggt ccggagaagg cgttccgtga actggcggac 660
gctctgctgg aaaaaggtaa cccggaaatg gttgacgttg ttacaggtgc gaccgtcagc 720
tctcagtctt tccgtcgtct gggtgctgct ctgctgcagt ctgctcgtcg tggtgaaaaa 780
gaagctatca tctctcgtgg ttctggtggt ggttctggtt ctggttctgg tggttctagt 840
catcacgaaa cccattacgg ttacgcgacc ctgagttacg cagattattg ggcaggcgaa 900
ctgggtcaaa gtcgcgacgt tctgctggca ggtaacgcag aagcagatcg cgcaggtgat 960
ctggacgcag gtatgtttga cgcagtttct cgtgcaaccc acggtcacgg cgcatttcgt 1020
caacagtttc agtacgcggt ggaagttctg ggcgaaaaag tgctgagcaa acaggaaacc 1080
gaagatagcc gcggtcgcaa aaaatgggaa tacgaaaccg acccgagcgt taccaaaatg 1140
gttcgcgcaa gcgcaagctt tcaagatctg ggcgaagacg gcgaaatcaa attcgaagca 1200
gtggaaggcg cagttgcact ggcagatcgc gcaagtagct ttatggtgga tagcgaagag 1260
tacaaaatca ccaacgtgaa agtgcacggc atgaaatttg ttccggttgc agttccgcac 1320
gaactgaaag gcatcgcgaa agagaaattc cattttgtgg aagatagccg cgtgaccgaa 1380
aacaccaacg gcctgaaaac catgctgacc gaagatagct ttagcgcgcg taaagtgagc 1440
agtatggaaa gtccgcacga tctggttgtt gataccgtag gtaccgttta ccactctcgt 1500
ttcggttctg acgctgaggc ttctgttatg ctgaaacgtg ctgacggttc tgaactgtct 1560
caccgtgagt tcatcgacta cgttatgaac ttcaacaccg ttcgttacga ctactacggt 1620
gacgacgctt cttacaccaa cctgatggct tcttacggta ccaaacactc tgctgactct 1680
tggtggaaaa ccggtcgtgt tccgcgtatc tcttgcggta tcaactacgg tttcgaccgt 1740
ttcaaaggtt ctggtccggg ttactaccgt ctgaccctga tcgctaacgg ttaccgtgac 1800
gttgttgctg acgttcgttt cctgccgaaa tacgaaggta acatcgacat cggtctgaaa 1860
ggtaaagttc tgaccatcgg tggtgctgac gctgaaaccc tgatggacgc tgctgttgac 1920
gttttcgctg acggtcagcc gaaactggtt tctgaccagg ctgtttctct gggtcagaac 1980
gttctgtctg ctgacttcac cccgggtacc gaatacaccg ttgaagttcg tttcaaagaa 2040
tttggttctg ttcgtgctaa agttgttgct caggctgctt acgctgctta cgtttcttgc 2100
accaccgttt gcccgcacgc tggtaaagct aaagctgaaa aagttgaagc tgctctgaaa 2160
ggtggtatct tccgtggtac cctgccggct gctgacgctc cgggtatcga caccaccgtt 2220
accttcaacg ctgacggtac cgctcagaaa gttgaactgg ctctggaaaa aaaatctgct 2280
ccgtctccgc tgacctaccg tggtacctgg atggttcgtg aagac 2325
<210> 27
<211> 775
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<400> 27
Val Ser Cys Thr Thr Val Cys Pro His Ala Gly Lys Ala Lys Ala Glu
1 5 10 15
Lys Val Glu Ala Ala Leu Lys Gly Gly Ile Phe Arg Gly Thr Leu Pro
20 25 30
Ala Ala Asp Ala Pro Gly Ile Asp Thr Thr Val Thr Phe Asn Ala Asp
35 40 45
Gly Thr Ala Gln Lys Val Glu Leu Ala Leu Glu Lys Lys Ser Ala Pro
50 55 60
Ser Pro Leu Thr Tyr Arg Gly Thr Trp Met Val Arg Glu Asp Gly Ile
65 70 75 80
Val Glu Leu Ser Leu Val Ser Ser Glu Gln Ser Lys Ala Pro His Glu
85 90 95
Lys Glu Leu Tyr Glu Leu Ile Asp Ser Asn Ser Val Arg Tyr Met Gly
100 105 110
Ala Pro Gly Ala Gly Lys Pro Ser Lys Glu Met Ala Pro Phe Tyr Val
115 120 125
Leu Lys Lys Thr Lys Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
130 135 140
Ser Ser Ser Ile Pro Asn Gly Thr Tyr Arg Ala Thr Tyr Gln Asp Phe
145 150 155 160
Asp Glu Asn Gly Trp Lys Asp Phe Leu Glu Val Thr Phe Asp Gly Gly
165 170 175
Lys Met Val Gln Val Val Tyr Asp Tyr Gln His Lys Glu Gly Arg Phe
180 185 190
Lys Ser Gln Asp Ala Asp Tyr His Arg Val Met Tyr Ala Ser Ser Gly
195 200 205
Ile Gly Pro Glu Lys Ala Phe Arg Glu Leu Ala Asp Ala Leu Leu Glu
210 215 220
Lys Gly Asn Pro Glu Met Val Asp Val Val Thr Gly Ala Thr Val Ser
225 230 235 240
Ser Gln Ser Phe Arg Arg Leu Gly Ala Ala Leu Leu Gln Ser Ala Arg
245 250 255
Arg Gly Glu Lys Glu Ala Ile Ile Ser Arg Gly Ser Gly Gly Gly Ser
260 265 270
Gly Ser Gly Ser Gly Gly Ser Ser His His Glu Thr His Tyr Gly Tyr
275 280 285
Ala Thr Leu Ser Tyr Ala Asp Tyr Trp Ala Gly Glu Leu Gly Gln Ser
290 295 300
Arg Asp Val Leu Leu Ala Gly Asn Ala Glu Ala Asp Arg Ala Gly Asp
305 310 315 320
Leu Asp Ala Gly Met Phe Asp Ala Val Ser Arg Ala Thr His Gly His
325 330 335
Gly Ala Phe Arg Gln Gln Phe Gln Tyr Ala Val Glu Val Leu Gly Glu
340 345 350
Lys Val Leu Ser Lys Gln Glu Thr Glu Asp Ser Arg Gly Arg Lys Lys
355 360 365
Trp Glu Tyr Glu Thr Asp Pro Ser Val Thr Lys Met Val Arg Ala Ser
370 375 380
Ala Ser Phe Gln Asp Leu Gly Glu Asp Gly Glu Ile Lys Phe Glu Ala
385 390 395 400
Val Glu Gly Ala Val Ala Leu Ala Asp Arg Ala Ser Ser Phe Met Val
405 410 415
Asp Ser Glu Glu Tyr Lys Ile Thr Asn Val Lys Val His Gly Met Lys
420 425 430
Phe Val Pro Val Ala Val Pro His Glu Leu Lys Gly Ile Ala Lys Glu
435 440 445
Lys Phe His Phe Val Glu Asp Ser Arg Val Thr Glu Asn Thr Asn Gly
450 455 460
Leu Lys Thr Met Leu Thr Glu Asp Ser Phe Ser Ala Arg Lys Val Ser
465 470 475 480
Ser Met Glu Ser Pro His Asp Leu Val Val Asp Thr Val Gly Thr Val
485 490 495
Tyr His Ser Arg Phe Gly Ser Asp Ala Glu Ala Ser Val Met Leu Lys
500 505 510
Arg Ala Asp Gly Ser Glu Leu Ser His Arg Glu Phe Ile Asp Tyr Val
515 520 525
Met Asn Phe Asn Thr Val Arg Tyr Asp Tyr Tyr Gly Asp Asp Ala Ser
530 535 540
Tyr Thr Asn Leu Met Ala Ser Tyr Gly Thr Lys His Ser Ala Asp Ser
545 550 555 560
Trp Trp Lys Thr Gly Arg Val Pro Arg Ile Ser Cys Gly Ile Asn Tyr
565 570 575
Gly Phe Asp Arg Phe Lys Gly Ser Gly Pro Gly Tyr Tyr Arg Leu Thr
580 585 590
Leu Ile Ala Asn Gly Tyr Arg Asp Val Val Ala Asp Val Arg Phe Leu
595 600 605
Pro Lys Tyr Glu Gly Asn Ile Asp Ile Gly Leu Lys Gly Lys Val Leu
610 615 620
Thr Ile Gly Gly Ala Asp Ala Glu Thr Leu Met Asp Ala Ala Val Asp
625 630 635 640
Val Phe Ala Asp Gly Gln Pro Lys Leu Val Ser Asp Gln Ala Val Ser
645 650 655
Leu Gly Gln Asn Val Leu Ser Ala Asp Phe Thr Pro Gly Thr Glu Tyr
660 665 670
Thr Val Glu Val Arg Phe Lys Glu Phe Gly Ser Val Arg Ala Lys Val
675 680 685
Val Ala Gln Ala Ala Tyr Ala Ala Tyr Val Ser Cys Thr Thr Val Cys
690 695 700
Pro His Ala Gly Lys Ala Lys Ala Glu Lys Val Glu Ala Ala Leu Lys
705 710 715 720
Gly Gly Ile Phe Arg Gly Thr Leu Pro Ala Ala Asp Ala Pro Gly Ile
725 730 735
Asp Thr Thr Val Thr Phe Asn Ala Asp Gly Thr Ala Gln Lys Val Glu
740 745 750
Leu Ala Leu Glu Lys Lys Ser Ala Pro Ser Pro Leu Thr Tyr Arg Gly
755 760 765
Thr Trp Met Val Arg Glu Asp
770 775

Claims (10)

1. A TP recombinant antigen comprising at least 1 TP17 antigen, at least 1 TP15 antigen, and at least 1 TP47 antigen; wherein,
the amino acid sequence of the TP17 antigen is as follows:
SEQ ID NO: 1; or
SEQ ID NO: 2;
the amino acid sequence of the TP15 antigen is as follows: SEQ ID NO: 3;
the amino acid sequence of the TP47 antigen is as follows:
SEQ ID NO: 4; or
SEQ ID NO: 5; or
SEQ ID NO: 6.
2. The TP recombinant antigen of claim 1, wherein the TP recombinant antigen comprises 1 TP17 antigen, 1 TP15 antigen, and 1 TP47 antigen.
3. The TP recombinant antigen of claim 1, wherein the TP recombinant antigen comprises 2TP 17 antigens, 1 TP15 antigen, and 1 TP47 antigen; wherein, the amino acid sequences of 2TP 17 antigens are all shown in SEQ ID NO: 1 or 2 of the amino acid sequences of the TP17 antigens are all shown as SEQ ID NO: 2, or 2 amino acid sequences of the TP17 antigens are shown as SEQ ID NOs: 1 and SEQ ID NO: 2, or a pharmaceutically acceptable salt thereof.
4. The TP recombinant antigen of claim 1, wherein the TP17 antigen, the TP15 antigen, and the TP47 antigen are linked directly or optionally through one or more linkers comprising (G) n, (GS) n, (SG) n, (GGGS) n, (ggggggs) n, or (AAY) n, wherein n is an integer of 1, 2, 3, 4, 5, or more.
5. The TP recombinant antigen according to claim 1, wherein the antigen comprises the following components in the order from N-terminal to C-terminal, TP17-TP15-TP47, TP15-TP17-TP47, TP17-TP15-TP47-TP17, or TP17-TP17-TP15-TP 47.
6. The TP recombinant antigen of any one of claims 1-5, wherein the TP recombinant antigen is a labeled antigen or a coating antigen; the N end of the TP recombinant antigen serving as a labeled antigen comprises a TRX label, and the N end of the TP recombinant antigen serving as a coating antigen comprises a GST label.
7. A nucleic acid encoding the TP recombinant antigen of any one of claims 1-6.
8. An expression vector comprising the nucleic acid of claim 7.
9. A host cell comprising the expression vector of claim 8, wherein the host cell comprises e.
10. A syphilis test strip, comprising the envelope antigen and the labeled antigen of claim 6, a TRX monoclonal antibody, and a label indirectly bound to the labeled antigen via the TRX monoclonal antibody.
CN202111523961.7A 2021-12-14 2021-12-14 TP recombinant antigen and preparation method and application thereof Pending CN114163539A (en)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102183646A (en) * 2010-12-16 2011-09-14 孙爱华 Preparation method of rTpN15-17-47-ELISA for detecting syphilis serum antibody
CN103298941A (en) * 2011-01-13 2013-09-11 奥索临床诊断有限公司 Treponema pallidum triplet antigen
CN111575308A (en) * 2020-05-25 2020-08-25 四川迈克生物新材料技术有限公司 Treponema pallidum recombinant chimeric antigen and preparation method and application thereof
CN113004380A (en) * 2021-02-18 2021-06-22 青岛硕景生物科技有限公司 Treponema pallidum recombinant antigen, preparation and application

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102183646A (en) * 2010-12-16 2011-09-14 孙爱华 Preparation method of rTpN15-17-47-ELISA for detecting syphilis serum antibody
CN103298941A (en) * 2011-01-13 2013-09-11 奥索临床诊断有限公司 Treponema pallidum triplet antigen
CN111575308A (en) * 2020-05-25 2020-08-25 四川迈克生物新材料技术有限公司 Treponema pallidum recombinant chimeric antigen and preparation method and application thereof
CN113004380A (en) * 2021-02-18 2021-06-22 青岛硕景生物科技有限公司 Treponema pallidum recombinant antigen, preparation and application

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Application publication date: 20220311